Nitric oxide (NO) is an important intercellular and intracellular messenger implicated in the pathogenesis of septic shock. Inhibition of NO synthase is under investigation as a treatment for hypotension in septic shock. In addition to the vasodilating effect of NO, this messenger also has effects on platelets and immune cells. In this investigation, we are examining the role of the NO pathway as a modulator of immune cell function and gene expression. We have been unable to create conditions under which human phagocytes, in particular neutrophils, endogenously produce NO (J Immunol: 1825, 1994). Therefore, the ability of NO produced by other cells, such as endothelium and epithelium, to alter the function of human phagocytes is being explored. We have confirmed that NO regulates cytokine production using a U937 monocytic cell line transfected to express murine inducible NO synthase (Blood: 1160, 1997). Further investigation of this effect has resulted in the description of a cGMP-independent signaling pathway for NO (J Biol Chem: 5959, 1997). We have found that in addition to upregulating TNFalpha production (J Immunol: 4102, 1994), NO modulates IL-8 message transcription and release in human neutrophil preparations. However, contrary to other reports, NO does not directly alter neutrophil chemotaxis (J Infect Dis: 116, 1998). More recent work has identified a NO-response element in the TNFalpha promoter (J Biol Chem, 1999). Recent experiments have generalized the role of this putative NO-response element to several unrelated promoters. Further, the importance of sequences flanking this NO-response element to its function are being investigated. Work with the IL-8 promoter suggest that this chemokine is regulated by NO through a mechanism that is also cGMP-independent, but distinct from the pathway that regulates TNFalpha. In a new phase of this project, expression microarrays are being used to define larger sets of genes regulated by NO and to dissect out the underlying mechanisms by which the regulation occurs.